One-dose vaccination associated with attenuated disease severity of adolescent and adult varicella cases in Beijing's Fengtai District.

BACKGROUND: In recent years, the number of varicella cases in adults has significantly increased in Beijing. However, the effect of the vaccination on varicella-related characteristics among adults has not been studied. METHODS AND RESULTS: Using data from the Infectious Disease Reporting System and the Immunization Information System, we compared the epidemiology and disease severity in breakthrough and unvaccinated varicella cases in adolescents and adults (≥ 15 year-old) from 2008 to 2011 in Beijing's Fengtai district, China. The results showed that the age (P = 0.003),contact history (90% vs. 73%, P = 0.019) and outbreak cases (10% vs. 1%, P < 0.0001) were significantly differently distributed between the two groups and that both the incidence of moderate-to-severe cases (26% vs. 45%, P = 0.035, OR = 0.446) and varicella-associated fever (49% vs. 66%, P = 0.068, OR = 0.534) were either significantly lower or trended to be lower in the breakthrough group than in the unvaccinated group. Additionally,vaccine effectiveness against moderate-to-severe cases of varicella was 55.4%. CONCLUSION: Altogether, these results indicate that vaccination against varicella among adolescents and adults affected the epidemiology and attenuated the disease severity of the cases. The Results from this study will provide useful information for the prevention of varicella in adolescents and adults.

High beta-cell mass prevents streptozotocin-induced diabetes in thioredoxin-interacting protein-deficient mice.

Thioredoxin-interacting protein (TxNIP) is an endogenous inhibitor of thioredoxin, a ubiquitous thiol oxidoreductase, that regulates cellular redox status. Diabetic mice exhibit increased expression of TxNIP in pancreatic islets, and recent studies suggest that TxNIP is a proapoptotic factor in beta-cells that may contribute to the development of diabetes. Here, we examined the role of TxNIP deficiency in vivo in the development of insulin-deficient diabetes and whether it impacted on pancreatic beta-cell mass and/or insulin secretion. TxNIP-deficient (Hcb-19/TxNIP(-/-)) mice had lower baseline glycemia, higher circulating insulin concentrations, and higher total pancreatic insulin content and beta-cell mass than control mice (C3H). Hcb-19/TxNIP(-/-) did not develop hyperglycemia when injected with standard multiple low doses of streptozotocin (STZ), in contrast to C3H controls. Surprisingly, although beta-cell mass remained higher in Hcb-19/TxNIP(-/-) mice compared with C3H after STZ exposure, the relative decrease induced by STZ was as great or even greater in the TxNIP-deficient animals. Consistently, cultured pancreatic INS-1 cells transfected with small-interfering RNA against TxNIP were more sensitive to cell death induced by direct exposure to STZ or to the combination of inflammatory cytokines interleukin-1beta, interferon-gamma, and tumor necrosis factor-alpha. Furthermore, when corrected for insulin content, isolated pancreatic islets from TxNIP(-/-) mice exhibited reduced glucose-induced insulin secretion. These data indicate that TxNIP functions as a regulator of beta-cell mass and influences insulin secretion. In conclusion, the relative resistance of TxNIP-deficient mice to STZ-induced diabetes appears to be because of an increase in beta-cell mass. However, TxNIP deficiency is associated with sensitization to STZ- and cytokine-induced beta-cell death, indicating complex regulatory roles of TxNIP under different physiological and pathological conditions.

Rescuing the subprime meltdown in insulin exocytosis in diabetes.

Neuroendocrine pancreatic islet beta-cells secrete the hormone insulin in response to glucose stimulation and adapt efficiently to increased demand by peripheral tissues to maintain glucose homeostasis. Insulin is packed within dense-core granules, which traffic and dock onto the plasma membrane whereby a Ca(2+) stimulus evokes exocytosis by soluble N-ethylmaleimide-sensitive factor attachment protein receptor (SNARE), complex-mediated, membrane fusion. Recent studies have unveiled postdocking steps mediated by "priming" factors that influence SNARE complex assembly to confer fusion readiness to the docked granules. This review will summarize recent insights into the priming role for Munc13 in the exocytosis of insulin granules. We present evidence for the interaction of Munc13-1 with exocytotic substrates involved in cAMP-mediated potentiation of insulin release, the latter we show to mediate enhanced granule-to-granule fusion events underlying compound exocytosis. We thus also further review the current understanding of granule-to-granule fusion. As agents acting on cAMP signaling are clinically used to augment insulin release in diabetes, this better understanding of priming steps may reveal additional novel therapeutic strategies to increase the capacity for insulin release to improve the treatment of diabetes.

The RalA GTPase is a central regulator of insulin exocytosis from pancreatic islet beta cells.

RalA is a small GTPase that is thought to facilitate exocytosis through its direct interaction with the mammalian exocyst complex. In this study, we report an essential role for RalA in regulated insulin secretion from pancreatic beta cells. We employed lentiviral-mediated delivery of RalA short hairpin RNAs to deplete endogenous RalA protein in mouse pancreatic islets and INS-1 beta cells. Perifusion of mouse islets depleted of RalA protein exhibited inhibition of both first and second phases of glucose-stimulated insulin secretion. Consistently, INS-1 cells depleted of RalA caused a severe inhibition of depolarization-induced insulin exocytosis determined by membrane capacitance, including a reduction in the size of the ready-releasable pool of insulin granules and a reduction in the subsequent mobilization and exocytosis of the reserve pool of granules. Collectively, these data suggest that RalA is a critical component in biphasic insulin release from pancreatic beta cells.

Activation of exchange protein directly activated by cyclic adenosine monophosphate and protein kinase A regulate common and distinct steps in promoting plasma membrane exocytic and granule-to-granule fusions in rat islet beta cells.

OBJECTIVES: Using FM1-43 epifluorescence imaging and electron microscopy, we recently reported that glucagon-like peptide (GLP-1)-mediated cyclic adenosine monophosphate (cAMP) potentiation of insulin secretion markedly promotes the number of plasma membrane (PM) exocytic sites and insulin secretory granule (SG)-to-granule fusions underlying compound and sequential exocytosis. METHODS: Here, we used FM1-43 imaging to dissect the distinct contributions of putative GLP-1/cAMP activated substrates--exchange protein directly activated by cAMP (EPAC) and protein kinase A (PKA)--in mediating these exocytic events. RESULTS: Like GLP-1, cAMP activation by forskolin increased the number of PM exocytic sites (2.3-fold), which were mainly of the robust-sustained (55.8%) and stepwise-multiphasic (37.7%) patterns corresponding to compound and sequential SG-SG exocytosis, respectively, with few monophasic hotspots (6.5%) corresponding to single-granule exocytosis. Direct activation of EPAC by 8-(4-chloro-phenylthio)-2'-O-methyladenosine-3',5'-cAMP also increased the number of exocytic sites, but which were mainly multiphasic (60%) and monophasic (40%) hotspots. Protein kinase A inhibition by H89 blocked forskolin-evoked robust-sustained hotspots, while retaining multiphasic (47%) and monophasic (53%) hotspots. Consistently, PKA activation (N6-benzoyladenosine-3',5'-cAMP) evoked only multiphasic (60%) and monophasic (40%) hotspots. These results suggested that PKA activation is required but alone is insufficient to promote compound SG-SG fusions. 8-(4-Chloro-phenylthio)-2'-O-methyladenosine-3',5'-cAMP plus N6-benzoyladenosine-3',5'-cAMP stimulation completely reconstituted the effects of forskolin, including increasing the number of exocytic sites, with a similar pattern of robust-sustained (42.6%) and stepwise (39.6%) hotspots and few monophasic (17.8%) hotspots. CONCLUSIONS: The EPAC and PKA modulate both distinct and common exocytic steps to potentiate insulin exocytosis where (a) EPAC activation mobilizes SGs to fuse at the PM, thereby increasing number of PM exocytic sites; and (b) PKA and EPAC activation synergistically modulate SG-SG fusions underlying compound and sequential exocytoses.

SNAREing voltage-gated K+ and ATP-sensitive K+ channels: tuning beta-cell excitability with syntaxin-1A and other exocytotic proteins.

The three SNARE (soluble N-ethylmaleimide-sensitive factor attachment protein receptor) proteins, syntaxin, SNAP25 (synaptosome-associated protein of 25 kDa), and synaptobrevin, constitute the minimal machinery for exocytosis in secretory cells such as neurons and neuroendocrine cells by forming a series of complexes prior to and during vesicle fusion. It was subsequently found that these SNARE proteins not only participate in vesicle fusion, but also tether with voltage-dependent Ca(2+) channels to form an excitosome that precisely regulates calcium entry at the site of exocytosis. In pancreatic islet beta-cells, ATP-sensitive K(+) (K(ATP)) channel closure by high ATP concentration leads to membrane depolarization, voltage-dependent Ca(2+) channel opening, and insulin secretion, whereas subsequent opening of voltage-gated K(+) (Kv) channels repolarizes the cell to terminate exocytosis. We have obtained evidence that syntaxin-1A physically interacts with Kv2.1 (the predominant Kv in beta-cells) and the sulfonylurea receptor subunit of beta-cell K(ATP) channel to modify their gating behaviors. A model has proposed that the conformational changes of syntaxin-1A during exocytosis induce distinct functional modulations of K(ATP) and Kv2.1 channels in a manner that optimally regulates cell excitability and insulin secretion. Other proteins involved in exocytosis, such as Munc-13, tomosyn, rab3a-interacting molecule, and guanyl nucleotide exchange factor II, have also been implicated in direct or indirect regulation of beta-cell ion channel activities and excitability. This review discusses this interesting aspect that exocytotic proteins not only promote secretion per se, but also fine-tune beta-cell excitability via modulation of ion channel gating.

Interaction between Munc13-1 and RIM is critical for glucagon-like peptide-1 mediated rescue of exocytotic defects in Munc13-1 deficient pancreatic beta-cells.

OBJECTIVE: Glucagon-like peptide-1 (GLP-1) rescues insulin secretory deficiency in type 2 diabetes partly via cAMP actions on exchange protein directly activated by cAMP (Epac2) and protein kinase A (PKA)-activated Rab3A-interacting molecule 2 (Rim2). We had reported that haplodeficient Munc13-1(+/-) mouse islet beta-cells exhibited reduced insulin secretion, causing glucose intolerance. Munc13-1 binds Epac2 and Rim2, but their functional interactions remain unclear. RESEARCH DESIGN AND METHODS: We used Munc13-1(+/-) islet beta-cells to examine the functional interactions between Munc13-1 and Epac2 and PKA. GLP-1 stimulation of Munc13-1(+/-) islets normalized the reduced biphasic insulin secretion by its actions on intact islet cAMP production and normal Epac2 and Rim2 levels. RESULTS: To determine which exocytotic steps caused by Munc13-1 deficiency are rescued by Epac2 and PKA, we used patch-clamp capacitance measurements, showing that 1) cAMP restored the reduced readily releasable pool (RRP) and partially restored refilling of a releasable pool of vesicles in Munc13-1(+/-) beta-cells, 2) Epac-selective agonist [8-(4-chloro-phenylthio)-2'-O-methyladenosine-3',5'-cyclic monophosphate] partially restored the reduced RRP and refilling of a releasable pool of vesicles, and 3) PKA blockade by H89 (leaving Epac intact) impaired cAMP ability to restore the RRP and refilling of a releasable pool of vesicles. Conversely, PKA-selective agonist (N(6)-benzoyladenosine-cAMP) completely restored RRP and partially restored refilling of a releasable pool of vesicles. To determine specific contributions within Epac-Rim2-Munc13-1 interaction sites accounting for cAMP rescue of exocytosis caused by Munc13-1 deficiency, we found that blockade of Rim2-Munc13-1 interaction with Rim-Munc13-1-binding domain peptide abolished cAMP rescue, whereas blockade of Epac-Rim2 interaction with Rim2-PDZ peptide only moderately reduced refilling with little effect on RRP. CONCLUSIONS: cAMP rescue of priming defects caused by Munc13-1 deficiency via Epac and PKA signaling pathways requires downstream Munc13-1-Rim2 interaction.

Munc13-1 deficiency reduces insulin secretion and causes abnormal glucose tolerance.

Munc13-1 is a diacylglycerol (DAG) receptor that is essential for synaptic vesicle priming. We recently showed that Munc13-1 is expressed in rodent and human islet beta-cells and that its levels are reduced in islets of type 2 diabetic humans and rat models, suggesting that Munc13-1 deficiency contributes to the abnormal insulin secretion in diabetes. To unequivocally demonstrate the role of Munc13-1 in insulin secretion, we studied heterozygous Munc13-1 knockout mice (+/-), which exhibited elevated glucose levels during intraperitoneal glucose tolerance tests with corresponding lower serum insulin levels. Munc13-1(+/-) mice exhibited normal insulin tolerance, indicating that a primary islet beta-cell secretory defect is the major cause of their hyperglycemia. Consistently, glucose-stimulated insulin secretion was reduced 50% in isolated Munc13-1(+/-) islets and was only partially rescued by phorbol ester potentiation. The corresponding alterations were minor in mice expressing one allele of a Munc13-1 mutant variant, which does not bind DAG (H567K/+). Capacitance measurements of Munc13-1(+/-) and Munc13-1(H567k/+) islet beta-cells revealed defects in granule priming, including the initial size and refilling of the releasable pools, which become accentuated by phorbol ester potentiation. We conclude that Munc13-1 plays an important role in glucose-stimulated insulin secretion and that Munc13-1 deficiency in the pancreatic islets as occurs in diabetes can reduce insulin secretion sufficient to cause abnormal glucose homeostasis.

Glucagon-like peptide 1 regulates sequential and compound exocytosis in pancreatic islet beta-cells.

Glucagon-like peptide 1 (GLP-1) has been postulated to potentiate insulin secretion by cAMP-mediated enhancement of mobilization and priming of secretory granules, but the precise exocytic events are unknown. We used epi-fluorescent microscopy of the fluorescent dye FM1-43, which incorporates into the plasma membrane and the exocytosing secretory granules (appearing as plasma membrane hotspots). KCl evoked exocytosis of 1.8 +/- 0.5 hotspots/rat beta-cell at the cell periphery, 82% of which are single transient increases of low amplitudes (151 +/- 7%), suggesting single secretory granule exocytosis; and the remaining 18% are stepwise increases in plasma membrane hotspots with higher amplitudes (170 +/- 9%), suggesting sequential secretory granule to secretory granule exocytic fusions. Addition of GLP-1 increased the hotspots to 6.0 +/- 0.7/beta-cell and exhibited a larger number of stepwise (41%) than transient (10%) increases with higher amplitudes of 259 +/- 19 and 278 +/- 23%, respectively. More interestingly, GLP-1 also evoked a robust and sustained pattern (49%) with even higher amplitudes of 354 +/- 18%, which are likely accelerated sequential secretory granule-secretory granule fusions. Electron microscopy studies collaborated with these imaging results, showing that GLP-1 increased the number of docked secretory granules at the plasma membrane and also increased the number of events showing direct contact of oncoming secretory granules with secretory granules undergoing exocytosis. We conclude that the potentiation of insulin secretion by GLP-1 is contributed by the mobilization of more insulin secretory granules to dock at the plasma membrane and the acceleration of sequential secretory granule-secretory granule fusions.